Incubation of human macrophages with human plasma apolipoprotein (Apo)A-I resulted in rounding of macrophages and decreased cell density that was independent of cell toxicity. Using a bromodeoxyuridine incorporation cell proliferation assay, we demonstrated that human plasma ApoA-I as low as 1 &#956;g/ml resulted in a >90% inhibition in macrophage proliferation. Using a recombinant version of ApoA-I, we found that the results were dependent on the expression system: ApoA-I expressed in human cells did not inhibit proliferation but ApoA-I expressed in Escherichia coli did. Human plasma-derived ApoA-II did not inhibit proliferation, but human plasma ApoA-IV did. We obtained all apolipoproteins from commercial sources. We considered the possibility that endotoxin contamination of the apolipoproteins contributed to the differential inhibition of macrophage cell proliferation. Endotoxin alone very potently inhibited macrophage proliferation (0.1 ng/ml inhibited macrophage proliferation >90%). Measurement of endotoxin levels in the apolipoprotein products, including an analysis of free versus total endotoxin that was masked because of binding to protein, suggested that endotoxin mediated the inhibition of macrophage proliferation. While human cell-derived recombinant ApoA-I and human plasma-derived ApoA-I showed similar levels of total endotoxin contamination (levels that were sufficient to inhibit macrophage proliferation), the level of free endotoxin in the human cell-derived recombinant ApoA-I was more than 1000-fold less than the ApoA-I level in human plasma-derived ApoA-I. This finding helps explain why the human cell-derived recombinant ApoA-I did not inhibit macrophage proliferation, but the plasma-derived ApoA-I did inhibit macrophage proliferation. Our findings show that endotoxin contamination can significantly influence apparent apolipoprotein-mediated cell effects (or effects of any other biological products), especially when these products are tested on highly endotoxin-sensitive cells, such as macrophages. Lastly, endotoxin potently inhibits human macrophage proliferation, which may be a significant pathogenic mechanism by which bacteria escape the immune system.